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Barriers in systemic delivery and preclinical testing of synthetic microRNAs in animal models: an experimental study
T Machackova1, P Vychytilova-Faltejskova, K Souckova
1Central European Institute of Technology (CEITEC), Masaryk University, Brno, Czech Republic. on.slaby@gmail.com.
Abstract:
Mus musculus is the most commonly used animal model in microRNA research; however, little is known about the endogenous miRNome of the animals used in the miRNA-targeting preclinical studies with the human xenografts. In the presented study, we evaluated the NOD/SCID gamma mouse model for the preclinical study of systemic miR-215-5p substitution with a semitelechelic poly[N-(2-hydroxypropyl)-methacrylamide]-based carrier conjugated with miR-215-5p-mimic via a reductively degradable disulfide bond. Murine mmu-miR-215-5p and human hsa-miR-215-5p have a high homology of mature sequences with only one nucleotide substitution. Due to the high homology of hsa-miR-215-5p and mmu-hsa-miR-215-5p, a similar expression in human and NOD/SCID gamma mice was expected. Expression of mmu-miR-215 in murine organs did not indicate tissue-specific expression and was highly expressed in all examined tissues. All animals included in the study showed a significantly higher concentration of miR-215-5p in the blood plasma compared to human blood plasma, where miR-215-5p is on the verge of a reliable detection limit. However, circulating mmu-miR-215-5p did not enter the human xenograft tumors generated with colorectal cancer cell lines since the levels of miR-215-5p in control tumors remained notably lower compared to those originally transfected with miR-215-5p. Finally, the systemic administration of polymer-miR-215-5p-mimic conjugate to the tail vein did not increase miR-215-5p in NOD/SCID gamma mouse blood plasma, organs, and subcutaneous tumors. It was impossible to distinguish hsa-miR-215-5p and mmu-miR-215-5p in the murine blood and organs due to the high expression of endogenous mmu-miR-215-5p. In conclusion, the examination of endogenous tissue and circulating miRNome of an experimental animal model of choice might be necessary for future miRNA studies focused on the systemic delivery of miRNA-based drugs conducted in the animal models.
Insights
Understanding the mouse miRNome is crucial for preclinical miRNA drug studies. High endogenous miR-215-5p in mice complicates systemic delivery research, hindering accurate assessment of human miR-215-5p substitution.
Area of Science:
- Biomedical Research
- Molecular Biology
- Pharmacology
Background:
- MicroRNA (miRNA) research frequently utilizes Mus musculus as an animal model.
- Limited knowledge exists regarding the endogenous miRNome in mice used for preclinical miRNA-targeting studies involving human xenografts.
- NOD/SCID gamma mice are a common model for human xenografts, but their endogenous miRNA profile requires characterization for miRNA therapeutics.
Purpose of the Study:
- To evaluate the NOD/SCID gamma mouse model for preclinical studies of systemic miR-215-5p substitution.
- To assess the feasibility of using a poly(N-(2-hydroxypropyl)-methacrylamide)-based carrier for delivering miR-215-5p mimics in mice.
- To investigate the endogenous expression of miR-215 in mice and its potential interference with human miR-215 studies.
Main Methods:
- Systemic administration of a polymer-conjugated miR-215-5p mimic via tail vein injection.
- Evaluation of miR-215-5p expression in blood plasma, organs, and subcutaneous xenograft tumors of NOD/SCID gamma mice.
- Comparison of endogenous mouse miR-215-5p levels with expected human miR-215-5p levels.
Main Results:
- Endogenous mmu-miR-215 expression was high and non-tissue-specific across all examined murine organs.
- Circulating miR-215-5p levels in mice were significantly higher than in human plasma.
- Systemic administration of the polymer-miR-215-5p conjugate did not lead to detectable increases in miR-215-5p in mouse plasma, organs, or tumors.
- High endogenous mmu-miR-215-5p levels prevented the distinction between mouse and human miR-215-5p in murine samples.
Conclusions:
- The high endogenous expression of mmu-miR-215-5p in NOD/SCID gamma mice complicates the assessment of systemic delivery of miR-215-5p mimics.
- It is challenging to differentiate between endogenous mouse miR-215-5p and administered human miR-215-5p due to sequence homology and high mouse expression.
- Characterizing the endogenous miRNome of experimental animal models is essential for future miRNA-based drug delivery studies.
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